Molecular Neurobiology (2019) 56:595–610 https://doi.org/10.1007/s12035-018-1065-1 Developmental Changes in Oligodendrocyte Genesis, Myelination, and Associated Behavioral Dysfunction in a Rat Model of Intra-generational Protein Malnutrition Nisha Patro1 & Aijaz Ahmad Naik1,2 & Ishan K. Patro1,2 Received: 20 February 2018 /Accepted: 5 April 2018 /Published online: 12 May 2018 # Springer Science+Business Media, LLC, part of Springer Nature 2018 Abstract Impairments in oligodendrocyte development and resultant myelination deficits appear as a common denominator to all neurological diseases. An optimal in utero environment is obligatory for normal fetal brain development and later life brain functioning. Late embryonic and early postnatal brains from F1 rat born to protein malnourished mothers were studied through a combination of immunocytochemical and quantitative PCR assay for analyzing the relative expression of platelet-derived growth factor receptor-α (PDGFRα), myelin-associated glycoprotein (MAG), proteolipid protein (PLP), and myelin oligodendrocyte glycoprotein (MOG) to determine oligodendrocyte genesis, differentiation, matura- tion, and myelination. Myelin integrity and corpus callosum caliber was assessed by Luxol fast blue (LFB) staining, whereas grip strength test and open field activity monitoring for behavioral evaluation in F1 rats. We demonstrate that intra-generational protein deprivation results in drastically low PDGFRα+ oligodendrocyte precursor (OPC) population and significantly reduced expression of myelin protein genes resulting in poor pre-myelinating and mature myelinating oligodendrocyte number, hypo-myelination, and misaligned myelinated fibers. LFB staining and MOG immunolabeling precisely revealed long-term changes in corpus callosum (CC) caliber and demyelination lesions in LP brain supporting the behavioral and cognitive changes at early adolescence and adulthood following maternal protein malnutrition (PMN). Thus, intra-generational PMN negatively affects the oligodendrocyte development and maturation resulting in myelination impairments and associated with behavioral deficits typically mimicking clinical hallmarks of neuropsychi- atric disorders. Our results further strengthen and augment the hypothesis BImpaired gliogenesis is a big hit for neuro- psychiatric phenotype.^ Keywords Maternal protein malnutrition . Oligodendrogenesis . Demyelination . Corpus callosum Introduction contribute to neural plasticity [1–3]. Earlier studies have iden- tified a number of markers for oligodendrocyte lineage includ- Myelination, essential for the propagation and speedy neuro- ing PDGFRα, NKX2.2, Olig-1, Olig-2, NG2, and even the transmission within the CNS, involves interfasicular oligo- domains in brain and spinal cord from where these cells arise dendrocytes. Oligodendrocytes, first characterized by del [4–6]. Oligodendrocytes in the CNS and Schwann cells in Rio-Hortega in 1928, not only serve functions of ensheathing the peripheral nervous system (PNS) synthesize myelin, a the axons for conduction of fast saltatory action potential but multi-layered, lipid-rich spiral sheath composed of sail-like also provide integrity and metabolic support to axons and extensions of plasma membrane around an axon, exclu- sively in vertebrates [5, 7, 8]. Myelin, a vital component of white matter, contributing ~ 40–50% in dry weight ex- pands radially and longitudinally almost simultaneously in * Ishan K. Patro sheaths. The continuous addition of lamellae from the [email protected] ab-axonal towards the ad-axonal side is followed by com- 1 School of Studies in Neuroscience, Jiwaji University, paction of multiple membrane layers, necessary for fast Gwalior 474011, India axonal conduction [2, 9]. Any deficit in the compaction 2 School of Studies in Zoology, Jiwaji University, Gwalior 474011, process may have negative consequences on both the trans- India mission of the signals and axonal health. 596 Mol Neurobiol (2019) 56:595–610 The formation of myelin from oligodendrocytes is a disrupted by environmental stressors leading to drastic prolonged process, continuing from childhood to adoles- changes in the myelination [24, 25]. Disruption of myelin cence in humans, where myelin build-up does continue and aberrant myelination as a result of malnutrition, bacte- into adulthood, though at a reduced rate [10, 11]. rial infection, inflammation, and toxicant exposure includ- Whereas, in rodent brain, events of myelination are ing ethanol and heavy metals, has also been well docu- programed predominantly postnatally, beginning at birth mented [26–29]. Recent study by Tilborg et al. [30] evi- in the spinal cord and achieved throughout the brain by denced delayed cortical myelination, oligodendrocyte matura- PND 45–60 [12, 13]. Myelination tends to proceed along tion, and autism-like behavior in rats following combined fetal with development of different brain areas. However, the inflammation with postnatal hypoxia. Although, earlier stud- start of myelination is not based on defined centers rather ies have reported neuropsychiatric symptoms in patients with occurring in response to functional demand, with brain multiple sclerosis, contusion-type spinal cord injury (SCI), areas related to early nursing, such as suckling reflex de- stroke, and metachromatic leukodystrophy [31, 32]; however, veloping first (birth to PND 10), followed by others, like what role oligodendrocytes and myelin play in higher brain motor and sensory regions (PND 10–24), and finally functions and neuropsychiatric conditions like schizophrenia learning areas (PND 17–37) in rats [12, 13]. Delayed re- and autism remains largely unclear [17]. Previous reports in flex maturation and other physical landmarks in F1 pups schizophrenia and bipolar patients have reported reduced den- born to protein-malnourished mothers, as we have previ- sity of oligodendrocytes in pre-frontal cortices, loss of white ouslyreported[14], incited us to investigate whether matter integrity, and increased microglial density in myelinat- intra-generational PMN affects the spatio-temporal oligo- ed fiber proximity along with abnormal connectivity in corpus dendrocyte genesis, turnover, and myelination status in callosum [33–36]. Autopsy studies of some mentally ill sub- the developing rat brain. jects have revealed subtle abnormalities in myelination as hall- Theroleofmyelininsensoryandmotorfunctionshas marks of disease and the reason could be that such subtle been well established; however, the grave neurological myelination changes may disturb the millisecond precision outcome following severe myelination impairments in leu- critical for the signal propagation in higher cortical networks kodystrophies further illustrates the pivotal role of myelin [18, 37, 38]. Recent clinical report from Chopra et al. [39] in normal brain functioning [2, 15–18]. A direct role of evidenced that more highly myelinated tracts are associated oligodendrocyte dysfunction in neuropsychiatric condi- with increased cognitive processing speed supporting role of tions is agreeable as the timing of vigorous oligodendro- myelination in cognition. These findings and other reports cyte maturation and myelin formation in developing brain [17, 40, 41] link myelination deficits with the psychiatric dis- coincides with the peak age for onset of clinical symptoms orders. However, whether the subtle changes in the white mat- of anxiety disorders [12, 19]. Besides their role in ter and mild myelination abnormalities result in gross behav- myelination, oligodendrocytes also provide trophic factors ioral changes or disease phenotype is yet poorly understood. and metabolites critical for neuronal function modulation Thus, it becomes worth to understand the impact of and other higher order brain functions [1, 20]. A myriad of intra-generational PMN on the genesis, density of OPCs, earlier studies support poor myelination and white matter and their differentiation and maturation along with dysfunction as a well-established component of large num- myelination across brain development. ber of neurodegenerative and neuropsychiatric disorders As it is increasingly clear that myelination is central to including multiple sclerosis (MS), vascular dementia, neuronal signaling finally contributing to a good cognitive schizophrenia, and Alzheimer’s[17, 21, 22]. Insults during outcome, dysfunction of cognitive abilities like learning, early brain development are considered as crucial memory, and attention thus coincide with the intermediate components/causatives for aberrant and abnormal phenotype of neuropsychiatric conditions. Further support neurodevelopment resulting in later life neurological dys- to this association is from the fact that the myelination function. Early-life protein malnutrition, a major stressor events in the brain occur postnatally and are completed involves intra-uterine growth restriction (IUGR) leading to at adulthood around the same time when the incidences of infants born small for gestational age (SGA) resulting in the psychotic disorders like schizophrenia and autism are perinatal mortality and postnatal morbidity [14, 23]. Not at peak. Therefore, whether, early-life protein deprivation only Bin utero period^ is critical owing to the large-scale induces significant deficits in oligodendrocyte formation, genesis and migration of neurons and glia, even the neo- myelination, and resultant behavioral profile needs imme- natal and early postnatal period represents an important diate attention. The present study